The overall goal of this NIDDK Mentored Research Scientist Development Award is to provide an integrated research and educational program for the applicant that will lead to an independent academic career in translational research. Robert J. Desnick, PhD, MD, Professor and Chair of Genetics and Genomic Sciences and Associate Dean for Genome- Based Research, and Ihor Lemischka, PhD, Professor of Gene and Cell Therapy and Director of the Black Family Stem Cell Institute, will serve as Mentor and Co-Mentor, respectively. This applicant will devote 95% of her effort to the career development program which includes: 1) undertaking the proposed laboratory research, 2) training in stem cell biology and iPS techniques, 3) participation in Genetics and Stem Cell Biology "Work in Progress" sessions, journal clubs and seminars, and attendance at national scientific meetings, and 4) bi-weekly meetings with Mentors. The specific aims of the proposed research are: 1) to investigate the pathogenesis of the life-threatening acute neurological attacks in Acute Intermittent Porphyria (AIP), the most common hepatic porphyria [due to deficient activity of hydroxymethylbilane synthase (HMB-synthase)], and 2) to evaluate the effectiveness of hepatocyte and bone marrow transplantation for AIP as proof-of-concept for future induced pluripotent stem (iPS) cell-based therapies for patients with AIP and the other acute hepatic porphyrias who have recurrent attacks. Towards this goal, Aim 1 will investigate disease pathogenesis in the previously generated T1/T2 AIP knock-down mice that have an "inducible" biochemical phenotype and in the newly generated AIP knock-in mice (R167Q+/+) that have a severe biochemical and clinical phenotype. Aim 2 will evaluate the effectiveness of primary hepatocyte transplantation and bone marrow transplantation in clearing the accumulated plasma and urinary porphyrin precursors, 4-aminolevulenic acid (ALA) and porphobilinogen (PBG) in the severely affected R167Q+/+ mice. These studies will determine the number of HMB-synthase competent cells necessary to clear the circulating porphyrin precursors and whether the clearance also down-regulates hepatic 5'- aminolevulinic acid synthase 1 (ALAS1) activity. Aim 3 will assess whether transplanting HMB-synthase overexpressing hepatocytes and bone marrow (transduced ex vivo with lentiviral vectors prior to transplantation) can effectively decrease the number of engrafted cells required to clear ALA and PBG accumulation in the R167Q+/+ mice. This is of particular importance, as it is a major challenge to achieve high levels of donor cell engraftment and proliferation for diseases such as AIP that lack hepatocellular damage and thus do not provide the donor cells a proliferative advantage. Based on the results of Aims 2 and 3, Aim 4 will apply the latest iPS technology to develop treatment for the AIP mice as a model for future applications in patients. The rationale for cell transplantation to prevent the acute attacks in AIP is supported by recent studies in our laboratory demonstrating that intraperitoneal injection of an AAV2/8 vector overexpressing HMB-synthase effectively and continuously prevented the biochemical induction of an acute attack in the T1/T2 mouse model of AIP (see Preliminary Results, Section C.3.). This integrated research and educational program, together with the outstanding research environment at Mount Sinai, should facilitate the applicant's transition to an academic career as an independent translational researcher. PUBLIC HEALTH RELEVANCE: The proposed research investigates whether transplantation of hepatocytes and bone marrow cells can treat Acute Intermittent Porphyria (AIP), a hepatic disorder resulting from the deficiency of an enzyme that synthesizes heme. Development of a safe and effective treatment should protect patients from the life-threatening acute neurological attacks of AIP and improve their quality of life.